Enhancing Transgene Expression from Recombinant AAV8 Vectors in Different Tissues Using Woodchuck Hepatitis Virus Post-Transcriptional Regulatory Element.
Enhancing Transgene Expression from Recombinant AAV8 Vectors in Different Tissues Using Woodchuck Hepatitis Virus Post-Transcriptional Regulatory Element.
复制标题
使用土拨鼠肝炎病毒转录后调控元件增强重组 AAV8 载体在不同组织中的转基因表达。
DOI：
10.7150/ijms.14152
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发表时间：
2016
影响因子：
3.6
通讯作者：
Yu X
中科院分区：
文献类型：
--
作者：
Wang L;Wang Z;Zhang F;Zhu R;Bi J;Wu J;Zhang H;Wu H;Kong W;Yu B;Yu X
腺相关病毒（AAV）载体已被广泛用于基因治疗和基因功能研究，因为强转基因表达是积极结果的先决条件。据报道，与其他血清型相比，AAV 8是用于肝、脑和肌肉转导的最有效的AAV血清型。然而，AAV 8介导的人肝细胞转导相当差，与小鼠肝细胞相比效率低约20倍。因此，我们应用土拨鼠肝炎病毒转录后调节元件（WPRE）来增强由组合启动子（CAG启动子）与CMV-IE增强子和鸡β-肌动蛋白启动子驱动的AAV 8介导的转基因表达，以获得更有效的病毒载体。在体外和体内评价了来自携带红色荧光蛋白（RFP）报告基因的重组AAV 8（rAAV 8）载体的转基因表达，有或没有WPRE。结果表明，WPRE改善了不同细胞系中AAV 8介导的RFP表达，在动物的肝脏、脑或肌肉中转基因表达明显增加。这项研究的结果将有助于大大减少必须注射的病毒颗粒的数量，以达到基因治疗中转基因表达的治疗水平。因此，这种剂量降低可能会降低与高剂量病毒载体相关的潜在风险。
Adeno-associated virus (AAV) vectors have been utilized extensively in gene therapy and gene function studies, as strong transgene expression is a prerequisite for positive outcomes. AAV8 was reported as the most efficient AAV serotype for transduction of the liver, brain and muscle compared with other serotypes. However, AAV8-mediated transduction of human hepatocytes is rather poor with approximately 20-fold lower efficiency compared with that of mouse hepatocytes. Therefore, we applied the woodchuck hepatitis virus post-transcriptional regulatory element (WPRE) to enhance AAV8-mediated transgene expression driven by a combination promoter (CAG promoter) with a CMV-IE enhancer and chicken beta-actin promoter for a more efficient viral vector. Transgene expression from recombinant AAV8 (rAAV8) vectors harboring a red fluorescent protein (RFP) reporter gene with or without WPRE were evaluated in vitro and in vivo. The results demonstrated that WPRE improved AAV8-mediated RFP expression in different cell lines with clear increases of transgene expression in the liver, brain or muscle of animals. The findings of this study will help to substantially reduce the quantity of viral particles that must be injected in order to reach a therapeutic level of transgene expression in gene therapy. Consequently, such dose reductions may lessen the potential risks associated with high doses of viral vectors.